Journal of Pharmacognosy and Phytochemistry 2021; 10(2): 96-103

E-ISSN: 2278-4136 P-ISSN: 2349-8234 www.phytojournal.com Identification of some constituents of JPP 2021; 10(2): 96-103 Received: 04-01-2021 guazumifolia Kunth () leaves from Accepted: 19-02-2021 Sucre state, Venezuela Haydelba D’Armas a. Facultad de Ciencias de la Ingeniería, Universidad Haydelba D’Armas, Victoria Vásquez, Shailili Moreno and Gabriel Ordaz Estatal de Milagro, Milagro, Provincia de Guayas, Ecuador Abstract b. Laboratorio de Productos Some constituents of Helicteres guazumifolia Kunth leaves extracts, which were obtained by maceration Naturales y Lípidos, Departamento de Química, in petroleum ether and methanol, were proposed to separate and identify. Chemical separation was Universidad de Oriente performed by column and thin layer chromatography, while identification was done by Gas Cumaná, estado Sucre, Chromatography-Mass Spectrometric analysis (GC-MS) and 1D-2D Nuclear Magnetic Resonance 1 13 1 1 Venezuela experiments ( H, C, DEPT-135°, HMQC, HMBC, COSY H- H). Several fatty constituents were isolates from non-polar fractions of this Helicteres specie, such as: methyl cis-13-docosenoate, 1- Victoria Vásquez docosanol, and decyl decanoate. Other constituents were identified by GC-MS as 2-dodecanone, methyl a. Laboratorio de Productos hexadecanoate, butyl (2-methylbutyl) phthalate, ethyl hexadecanoate, n-eicosane, di (2-ethylhexyl) Naturales y Lípidos, phthalate, (22E, 24R)-stigmasta-4,22-dien-3-one, δ-stigmast-4-en-3-one, benzophenone, (4- Departamento de Química, methylphenyl) phenylmethanone, 6,10,14-trimethylpentadeca-2-one, dibutylphtalate, 4,8,12,16- Universidad de Oriente tetramethylheptadecan-4-olide, and cyclic 1,2-ethanediyl acetal (5α)-4,4-dimethyl-cholestan-3-one. Cumaná, estado Sucre, Results suggest that H. guazumifolia Kunth could biosynthesize several chemical families, including fatty Venezuela acid, hydrocarbon, terpenoid, steroid, and phenolic derivatives, which could suggest its possible b. Unidad Educativa Instituto ethnomedical uses. Libertador, Cumaná, estado Sucre, Venezuela Keywords: Helicteres, GC-MS, lipids, isoprenoids, NMR Shailili Moreno Laboratorio de Productos Introduction Naturales y Lípidos, Species of the Helicteres (Malvaceae, previously included in family Sterculaceae) are Departamento de Química, characterized by having distinctive fruits, which are spiral capsules composed of five Universidad de Oriente Cumaná, estado Sucre, unilocular carpels and may be ovoid to ellipsoid or subcylindrical (Golberg, 2009; Cowie, [1, 2] Venezuela 2011) . They are widely distributed around the world and their pharmacological potential has gained prominence, especially with H. isora and H. angustifolia that have a long history of Gabriel Ordaz use in traditional Chinese medicine; while about 149 compounds have been isolated from Laboratorio de Productos several of Helicteres species, including terpenoids, sterols, and phenolic compounds, among Naturales y Lípidos, [3] Departamento de Química, others (Fernandes et al., 2020) . Universidad de Oriente The specie H. guazumifolia Kunth is particularly abundant in several regions of America, such Cumaná, estado Sucre, as Mexico (Bravo et al., 2016; Notario et al., 2020) [4, 5], Costa Rica (Goldberg, 2009) [1], Venezuela Brazil (Fernandes and Oliveira, 2018; Stavis et al., 2020) [6, 7], Colombia (Angarita et al., 2014; Sanmartín-Sierra et al., 2016) [8, 9], and Venezuela (Lárez, 2007; Rondón and Cumana- [10-13] Campos, 2007; Fariñas et al., 2011, Díaz and Carrasco, 2014) . Population of these [14] regions seems to use this specie as medicinal (da Costa et al., 2020) , and although its ethno-botanical uses have not been indicated, they may be related to its effects on fertility and women’s health (Yazbek et al., 2016) [15]. Previous study of extracts obtained from aerial parts of H. guazumifolia Kunth showed slight

antimicrobial and antifungal activities, and strong toxic activity against Artemia salina [16] (D’Armas et al., 2020) . Furthemore, it has been reported the chemical compositions of its leaves essential oil, which was constituted mainly by diisobuthylphtalate, pentadecanal, 2- Corresponding Author: chloroethyl linoleate, hexahydrofarnesyl acetone, and isophytol, among others (Ordaz et al., Haydelba D’Armas 2011) [17]. For that, the aim of this study was to isolated and characterize some phytochemical a. Facultad de Ciencias de la Ingeniería, Universidad constituents from petroleum ether and methanol extracts of H. guazumifolia Kunth leaves Estatal de Milagro, Milagro, collected in Sucre state, Venezuela. Provincia de Guayas, Ecuador b. Laboratorio de Productos Materials and Methods Naturales y Lípidos, Collection and identification of plant materials Departamento de Química, Universidad de Oriente Sample of H. guazumifolia was collected in the way between Cumaná city and San Juan de Cumaná, estado Sucre, Macarapana sector (10°38’44’’N, 63°02’20’’W; 43 mamsl), Sucre state, Venezuela. Venezuela ~ 96 ~ Journal of Pharmacognosy and Phytochemistry http://www.phytojournal.com

Taxonomic identification was realized at the herbarium Identification by GC-MS was made by comparison of “Isidro Ramón Bermúdez Romero”, Biology Department, obtained values with those of WILEY and NIST databases, Universidad de Oriente, Sucre Campus, Venezuela. while assignations of chemical shift on NMR experiments were made by comparison with theoretical 1H and 13C NMR Extracts spectrums at Spectral Database for Organic Compounds Samples of dried and powdered H. guazumifolia Kunth leaves (AIST, 2017) [24], and NMR predictor available on-line were extracted with petroleum ether. Then, solvent was (Aires-de-Sousa et al., 2002; Steinbeck et al., 2003; Binev separated and evaporated under vacuum in a rotary evaporator and Aires-de-Sousa, 2004; Binev et al., 2004; Banfi and Heidolph (~11 mbar, 40 °C), obtaining crude petroleum ether Patiny, 2008; Castillo et al., 2011) [19-24]. extract (PEE). The vegetal residues were re-extracted with methanol. Solvent was separated and anhydrous sodium Results sulfate was added to dry (~5 g/100 mL of solvent). Then, Separation and analysis of fractions from PEE filtrated solvent was concentrated in the same conditions to According to yield and TLC analysis of the fractions from obtain crude methanol extract (ME). PEE, E2 and E3 were selected to be partitioned. Fraction E2 (157.9 mg) was chromatographed in column (silica gel 35-70 Separation mesh; petroleum ether-dicloromethane-ethyl acetate: 1:0:0, PEE of H. guazumifolia Kunth leaves (5.36 g) was 4:1:0, 3:2:0, 1:1:0, 2:3:0, 1:4:0, 0:1:0, 0:7:3, 0:1:1, 0:3:7, chromatographed in column (CC), using silica gel 35-70 mesh 0:0:1), obtaining 5 fractions (E2.1-E2.5, 96.39% w/w). as stationary phase in a relation in mass of 30:1 respect to Fraction E2.2 (58.6 mg) was separated by CC (silica gel 37- extract. Mobile phase was performed on basis to increasing 70 mesh, petroleum ether-dicloromethene: 1:0, 4:1, 3:2, 1:1, polarity with mixes of solvents in a relation in volume, 2:3, 1:4, 0:1) in 5 fractions (E2.2.1-E2.2.5, 87.37% w/w), starting with petroleum ether-dichloromethane (1:0, 9:1, 4:1, from which E2.2.5 (15.3 mg) was chromatographed by TLC 7:3, 3:2, 1:1, 2:3, 3:7, 1:4, 1:9, 0:1), then dichloromethane- (silica gel 60 mesh, petroleum ether-dicloromethene: 2:3), ethyl acetate (1:0, 4:1, 3:2, 1:1, 2:3, 1:4, 0:1), and finally ethyl obtaining 4 new fractions (E2.2.5.1-E2.2.5.4, 73.46% w/w). acetate-methanol (1:0, 4:1, 3:2, 1:1, 2:3, 1:4, 0:1). They were Fraction E2.2.5.2 (5.3 mg) was a white solid and showed to obtained 124 elutes, which were grouped in 15 fractions (E1- be a pure compound according TLC analysis (Rf = 0.26), E15, 2.35 g, 43.77% w/w). Thin layer chromatography (TLC) reason for which it was analyzed by NMR. on glass plates (20×20 cm2) covered with silica gel 60 mesh Chemical shifts are shown in table 1, which are consistent (0.5 mm) was used to join elutes according to separation with an unsaturated fatty acid methyl ester. The 13C-NMR observed under ultraviolet light and with ammonium spectrum of fraction E2.2.5.2 showed 15 signals, δC 174.30 molibdate solution (5% w/v) in aqueous H2SO4 (5% v/v). ppm corresponded to a quaternary carbon of the carbonyl Chromatography of ME of H. guazumifolia Kunth leaves group (C=O), δC 130.04-129.79 ppm were signals assigned to (5.08 g) was performed similarly with mixes of petroleum nucleus of a C=C bond (methynes, according to DEPT-135°), ether-ethyl acetate-methanol in a relation in volume of 1:0:0, δC 51.42 ppm was assigned to the primary carbon of the 4:1:0, 3:2:0, 1:1:0, 2:3:0, 1:4:0, 0:1:0, 0:4:1, 0:3:2, 0:1:1, methoxy group (-OCH3). Rocking methylene signals were 0:2:3, 0:1:4, and 0:0:1. It yielded 101 elutes, which were observed between δC 34.17 ppm and δC 22.72 ppm, and the grouped in 10 fractions (M1-M10, 3.16 g, 62.24% w/w). terminal methyl group appeared at δC 14.12 ppm of the 1 Continuous separation was performed by CC and preparative spectrum. Signals at δH 5.38-5.28 ppm (m) on the H-NMR TLC (1.0 mm of silica gel thickness). spectrum were assigned to the unsaturated protons, while the signal at δH 3.65 ppm (s) was attributed to methoxy protons. Characterization Chemical shifts around δH 2.28 ppm (t), δH 1.98 ppm (d), δH Some fractions obtained after a continuous chromatographic 1.60 ppm (s), δH 1.25 ppm (m) and δH 0.86 ppm (t), separation were analyzed by Infrared Spectroscopic (using a correspond to α-carbonyl methylene protons, α-methylene FTIR 16 PC spectrometer Perkin Elmer), Gas protons respect to double bond (α-CH2), β-carbonyl Chromatography-Mass Spectrometry (GC-MS, in a methylene protons, methylene protons of chain, and the chromatograph Hewlett Packard 5890 II with EI 70 eV, terminal methyl protons, respectively. Comparison of these column of methylsilicone of 25 m×0.18 DI×0.18 mm experimental spectral data with theoretical spectrums for thickness, T (injector) = 280 °C, Ti (oven) = 70°C, rate of several monounsaturated fatty acid methyl esters available at [24] 10°C/min, and Tf (oven) = 300 °C; coupled with a mass AIST (2017) led us to establish methyl cis-13-docosenoate spectrometer Hewlett Packard 5971 A; total running time was (methyl erucate, figure 1) as the compound isolated in fraction 45 min.), and 1D (1H, 13C) and 2D (HMBC, HMQC, COSY E2.2.5.2. 1H-1H) Nuclear Magnetic Resonance (NMR) (using spectrometers Bruker 600 MHz and Varian 500 MHz). 1H and 13C spectrum were made at 500.13 MHz and 125.75 MHz, respectively, in solution of deuterated chloroform (CDCl3), expressing chemical shifts in ppm respect to tetramethylsilane Fig 1: Chemical structure of methyl cis-13-docosenoate, possible (TMS). compound isolated as fraction E2.2.5.2 from petroleum ether extract of H. guazumifolia Kunth leaves

Table 1: Chemical shifts of NMR (1H and 13C) spectrums of fraction E2.2.5.2 obtained from petroleum ether extract of H. guazumifolia kunth leaves

a 1 1 a δC (ppm) DEPT 135° δH (ppm) COSY H- H 174.30 -(C=O)- (C1) - 130.04 * -CH=CH- (C13, C14) 5.34-5.30 (m) H12, H15 129.79 *

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51.42 -OCH3 (C23) 3.65 34.17 α-CH2 to C=O (C2) 2.31-2.25 (t) H3 31.95 β-CH2 to CH3 (C20) * 29.82 -(CH2)n- (C7, C16) * 29.73 -(CH2)n- (C7, C8, C9) * H13, H14 29.57 -(CH2)n- (C6, C18) 1.28-1.23 (m) * 29.37 -(CH2)n- (C5, C10, C17, C19) 29.19 γ-CH2 to C=O (C4) H3 22.71 α-CH2 to CH3 (C21) H22 27.27 α-CH2 to C=C (C12, C15) 2.00-1.98 (d) H13, H14 25.01 β-CH2 to C=O (C3) 1.59 (s) H2, H4 14.12 -CH3 (C22) 0.88-0.83 (t) H21 *Interchangeable. a Assignations were made on basis to theoretical spectrums of methyl cis-13-docosenoate structure (figure 1), SDBS No. 7657 (AIST, 2017) [24].

+· +· +· Fraction E2.3 was analyzed by GC-MS, identifying six 127 [M-C11H23] , 113 [M-C12H25] , 99 [M-C13H27] , 85 [M- +· +· +· majority compounds (figure 2): 2-dodecanone (C12H24O C14H29] , 71 [M-C15H31] , 57 [M-C16H33] base/main peak, -1 +· +· [calculated 184.3184 g mol ]; RT = 19.55 min, Area = 43 [M-C17H35] , 29 [M-C18H37] ), and di(2- +· +· 8.84%, Match = 94%; m/z = 184 [M] , 85 [M-C7H15] , 71 ethylhexyl)phthalate (DEHP, C24H38O4 [calculated 390.5561 +· +· -1 [M-C8H17] , 58 [M-C9H18] base/main peak, 43 [M- g mol ]; RT = 29.06 min, Area = 28.90%, Match = 91%; m/z +· +· +· +· C10H21] ), methyl hexadecanoate (C17H34O2 [calculated = 390 [M] , 279 [M-C8H15] , 167 [M-C12H15O4] , 149 [M- -1 +· +· 270.4507 g mol ]; RT = 20.71 min, Area = 15.52%, Match = C16H34O] base/main peak, 57 [M-C20H29O4] , 43 [M- +· +· +· +· 98%; m/z = 270 [M] , 239 [M-CH3O] , 227 [M-C3H7] , 199 C21H31O4] ). Furthermore, the IR spectrum showed signals +· +· +· -1 -1 [M-C5H11] , 185 [M-C6H13] , 171 [M-C7H15] , 129 [M- due to O-H (3308 cm ), C-H (2900-2890 cm ), C=O (1735 +· +· +· -1 -1 -1 C10H21] , 87 [M-C13H27] , 74 [M-C14H28] base/main peak, cm ), C-O (1172 cm ), and C-H (730 cm ) bonds. +· +· 57 [M-C13H25O2] , 43 [M-C14H27O2] ), butyl(2-methylbutyl) Separation of fraction E3 (100.0 mg) was made by CC (silica -1 phtalate (C17H24O4 [calculated 292.3701 g mol ]; RT = 21.43 gel 35-50 mesh, petroleum ether-ethyl acetate-methanol: min, Area = 23.83%, Match = 96%; m/z = 292 [M]+·, 223 [M- 1:0:0, 4:1:0, 3:2:0, 1:1:0, 2:3:0, 1:4:0, 0:1:0, 0:7:3, 0:1:1, +· +· +· C5H9] , 205 [M-C5H11O] , 149 [M-C9H19O] base/main 0:3:7, 0:0:1), yielding 5 fractions (E3.1-E3.5, 50.10% w/w). +· peak, 57 [M-C13H15O4] ), ethyl hexadecanoate (C18H36O2 Fraction E3.2 (11.6 mg, beige solid) was separated by CC [calculated 284.4772 g mol-1]; RT = 21.70 min, Area = (silica gel 35-50 mesh, petroleum ether-ethyl acetate: 1:0, 4:1, +· +· 13.11%, Match = 98%; m/z = 284 [M] , 241 [M-C3H7] , 213 3:2, 1:1, 2:3, 1:4, 0:1) in 4 new fractions (E3.2.1-E3.2.4, +· +· +· [M-C5H11] , 199 [M-C6H13] , 185 [M-C7H15] , 143 [M- 55.78% w/w). Fraction E3.2.4 (4.4 mg) was purified by +· +· +· C10H21] , 129 [M-C11H23] , 115 [M-C12H25] , 88 [M- preparative TLC (silica gel 60 mesh, petroleum ether-ethyl +· C14H28] base/main peak), n-eicosane (C20H42 [calculated acetate: 7:3), recovering a possible pure compound (fraction -1 282.5475 g mol ]; RT = 21.79 min, Area = 5.98%, Match = E3.2.4.1, 3.9 mg, 88.64% w/w, Rf = 0.70). This fraction was +· +· +· 1 13 95%; m/z = 282 [M] , 155 [M-C9H19] , 141 [M-C10H21] , analyzed by H and C NMR.

Fig 2: Chromatogram of fraction E2.3, pointing the main constituents. ~ 98 ~ Journal of Pharmacognosy and Phytochemistry http://www.phytojournal.com

Experimental spectral data of the fraction E3.2.4.1 is shown in and their comparison with data of theoretical and predicted table 2. The 13C-NMR spectrum showed 9 signals that almost spectrums for several long chain alcohols, the compound all (δC 63.16-22.71 ppm) correspond to methylene nucleuses isolated in fraction E3.2.4.1 could be 1-docosanol (figure 3). according to DEPT-135° experiment, except δC 14.11 ppm, which corresponds to terminal methyl (-CH3). Signals at δH OH 3.63 ppm (t) and 1.55 ppm (m) of oxygenated methylene protons and neighbor methylene protons, respectively Fig 3: Chemical structure of 1-docosanol, possible compound 1 appeared in H-NMR spectrum; δH 1.24 ppm (m) of other isolated as fraction E3.2.4.1 from petroleum ether extract of H. methylene protons, and δH 0.87 ppm (t) of terminal methyl guazumifolia Kunth leaves protons. These signals were consistent with a fatty alcohol,

Table 2: Chemical shifts of NMR (1H and 13C) spectrums of fraction E3.2.4.1 obtained from petroleum ether extract of H. guazumifolia kunth leaves

a 1 1 a δC (ppm) DEPT 135° δH (ppm) COSY H- H 63.16 α-CH2 to OH (C1) 3.65-3.60 (t) H2 32.91 β-CH2 to OH (C2) 1.55-1.53 (m) H1 31.97 β-CH2 to CH3 (C20) 29.72 -(CH2)n- (C5-C18) 29.48 δ-CH2 to OH (C4) 1.24 (m) 29.39 γ-CH2 to CH3 (C19) 25.81 γ-CH2 to OH (C3) 22.71 α-CH2 to CH3 (C21) H12 14.11 -CH3 (C22) 0.89-0.84 (t) H11 - - 1.58 (H-O) a Assignations were made on basis to theoretical spectrums of 1-docosanol (figure 3), SDBS No. 7647 (AIST, 2017) [24].

Fraction E3.4 was also purified by preparative TLC (silica gel long chain alcohol). According to experimental and 60 mesh, petroleum ether-dicloromethane: 3:2). Fraction theoretical data, the component isolated in fraction E3.4.1 recovered was other possible pure compound (E3.4.1, 3.4 mg, could be decyl decanoate (figure 4). 60.63%, Rf = 0.69) and was analyzed by NMR (table 3). The NMR experiments indicated a carbonyl nucleus (δC 173.94 O ppm), an oxygenated methylene (δC 63.38 ppm; δH 4.03 ppm), α-CH2 to carbonyl (δC 34.41 ppm; δH 2.27 ppm), β-CH2 to O O bridge (δC 28.64 ppm; δH 1.59 ppm), other saturated Fig 4: Chemical structure of decyl decanoate, possible compound methylenes (δC 31.91-25.02 ppm; δH 1.23 ppm), and terminal isolated as fraction E3.4.1 from petroleum ether extract of H. methyl groups (δC 14.10 ppm; δH 0.86 ppm). These signals are guazumifolia Kunth leaves consistent with a wax structure (ester of a fatty acid with a

Table 3: Chemical shifts of NMR (1H and 13C) spectrums of fraction E3.4.1 obtained from petroleum ether extract of H. guazumifolia Kunth leaves

a a δC (ppm) DEPT 135° δH (ppm), HMQC HMBC 173.94 -(C=O)- (C1) - - 64.38 -CH2O- (C11) 4.05-4.02 (t) H12 34.41 α-CH2 to C=O (C2) 2.28-2.25 (t) - 28.64 α-CH2 to -CH2O- (C12) 1.61-1.56 (m) H11, H13-H15 31.91 β-CH2 to CH3 (C8, C18) * H4-H10, 29.64 γ-CH2 to CH3 (C7) * H14-H20 29.52 γ-CH2 to CH3 (C17) * 29.46 γ-CH2 to C=O (C4) H2, H3, H5-H7 * 29.34 γ-CH2 to –CH2O- (C14) H13, H15-H17 * 1.23 (m) 29.26 -(CH2)n- (C5-C6) H3-H9 * 29.15 -(CH2)n- (C15-C16) H13-H19 25.93 β-CH2 to –CH2O- (C13) H11 25.02 β-CH2 to C=O (C3) H2 22.67 α-CH2 to CH3 (C9, C19) H6-H8, H10, H16-H18, H20 14.10 -CH3 (C10, C20) 0.87-0.85 (t) - *Interchangeable. a Assignations were made on basis to theoretical spectrums of decyl decanoate (figure 4), SDBS No. 7580 (AIST, 2017) [24].

+· +· +· Analysis of fractions from ME m/z = 410 [M] , 367 [M-C3H7] , 298 [M-C8H16] , 271 [M- +· +· +· According to TLC analyses, fractions M2 (7.9 mg) and M3 C10H19] , 147 [M-C18H31O] , 123 [M-C21H35] , 95 [M- +· +· +· (18.2 mg) seemed to contain a few compounds, which showed C22H35O] , 81 [M-C24H41] , 69 [M-C25H41O] , 55 [M- +· +· very close Rf. Due to their low mass, both fractions were C26H43] base/main peak, 43 [M-C26H39O] ), and δ-stigmast- - selected to be studied by GC-MS. Chromatogram of fraction 4-en-3-one (sitostenone, C29H48O [calculated 412.6908 g mol 1 +· +· M2 (figure 5) displayed two main peaks at 29.75 min ]; Match = 99%; m/z = 412 [M] , 370 [M-C3H6] , 281 [M- +· +· +· (39.28%) and 32.70 min (60.72%), which corresponded to C8H19O] , 207 [M-C14H21O] , 124 [M-C20H32O] base/main +· two steroidal constituents, (22E, 24R)-stigmasta-4,22-dien-3- peak, 43 [M-C26H41O] ), respectively. -1 one (C29H46O [calculated 410.6749 g mol ]; Match = 97%;

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Fig 5: Chromatogram of fraction M2, pointing the main constituents

Chromatogram of fraction M3 (figure 6) displayed 6 main peak), dibutylphtalate (DBP, C16H22O4 [calculated 278.3435 g peaks, related to the compounds benzophenone mol-1]; RT = 21.10 min, Area = 7.43%, Match = 96%; m/z = -1 +· +· +· (diphenylmethanone, C13H10O [calculated 182.2179 g mol ]; 278 [M] , 223 [M-C4H7] , 205 [M-C5H13] , 149 [M- +· +· RT = 16.13 min, Area = 15.40%, Match = 96%; m/z = 182 C8H17O] base/main peak, 57 [M-C12H13O4] ), 4,8,12,16- +· +· +· [M] , 105 [M-C6H5] base/main peak, 77 [M-C7H5O] , 51 tetramethylheptadecan-4-olide (5-methyl-5-(4,8,12- +· [M-C9H7O] ), 4-methylbenzophenone ((4- trimethyltridecyl)dihydrofuran-2(3H)-one, C21H40O2 -1 methylphenyl)phenylmethanone, C14H12O [calculated [calculated 324.5411 g mol ]; RT = 26.34 min, Area = -1 +· +· 196.2445 g mol ]; RT = 18.25 min, Area = 7.71%, Match = 10.58%, Match = 99%; m/z = 324 [M] , 254 [M-C5H10] , 184 +· +· +· +· +· +· 97%; m/z = 196 [M] , 181 [M-CH3] , 119 [M-C6H5] [M-C10H20] , 114 [M-C15H30] , 99 [M-C16H33] base/main +· +· +· +· +· base/main peak, 105 [M-C7H7] , 91 [M-C7H5O] , 77 [M- peak, 83 [M-C17H37] , 69 [M-C18H39] , 55 [M-C18H37O] ), +· +· +· C8H7O] , 65 [M-C9H7O] , 51 [M-C10H9O] ), 6,10,14- and cyclic 1,2-ethanediyl acetal (5α)-4,4-dimethyl-cholestan- -1 trimethylpentadeca-2-one (hexahydrofarnesyl acetone 3-one (C31H54O2 [calculated 458.7593 g mol ]; RT = 28.62 -1 +· C18H36O [calculated 268.4778 g mol ]; RT = 19.31 min, Area min, Area = 6.67%, Match = 90%; m/z = 430 [M-C2H4] , 340 +· +· +· +· +· = 43.33%, Match = 95%; m/z = 268 [M] , 225 [M-C3H7] , [M- C6H14O2] , 125 [M- C22H37O2] , 99 [M- C26H47] +· +· +· +· 210 [M-C3H6O] , 194 [M-C4H10O] , 85 [M-C13H27] , 71 base/main peak, 55 [M-C27H47O2] ). +· +· +· [M-C14H29] , 58 [M-C15H30] , 43 [M-C16H33] base/main

Fig 6: Chromatogram of fraction M3, pointing the main constituents ~ 100 ~ Journal of Pharmacognosy and Phytochemistry http://www.phytojournal.com

Discussion the compounds isolated and identified in this study are known Three lipid derivatives were isolated from H. guazumifolia for their biological activities. In this sense, this specie could Kunth petroleum ether extract, which were characterized by represent an important ethnobotanical resource. NMR as methyl cis-13-docosenoate, 1-docosanol and decyl decanoate, respectively. Methyl cis-13-docosenoate (or Acknowledgments methyl erucate) has been reported in the petroleum ether The authors want to thank Universidad Politécnica Territorial extract of Luffa echinata Roxb fruit (Rachana et al., 2019) [25], del Oeste de Sucre, Ecology Laboratory at Universidad Simón and bioactive oils of Horsfieldia glabra seeds (Waman et al., Bolívar; and both Instituto Venezolano de Investigaciones 2021) [26], and the Brassica nigra seed (Olgun et al., 2017) Científicas and Universidad Central de Venezuela, for IR, [27]. Fatty acid methyl esters from vegetable oils have shown GC-MS and NMR analyses, respectively. Also, we thank the antimicrobial and antioxidant activities, possibly due to the partial financial support that was made by Universidad de presence of unsaturated ones (Pinto et al., 2017) [28]. Oriente, and FUNDACITE (Sucre). The fatty alcohol 1-dodecanol is well known by its antiviral bioactivity against herpes simplex viruses 1 and 2 (Pope et al., References 1996; Leung and Sacks, 2005) [29, 30]. It has been isolated from 1. Goldberg L. Patterns of nectar production and the bark of Rhamnus caroliniana chloroform extract (Mekala composition, and morphology of floral nectaries in et al., 2017) [31], and identified in extracts of Kielmeyera Helicteres guazumifolia and Helicteres baruensis Coriacea leaf (Figueiredo et al., 2014) [32] and Gomphrena (): two sympatric species from the Costa decumbens Jacq. (Yamuna et al., 2017) [33]. While decyl Rican tropical dry forest. Rev. biol. Trop decanoate has been identified as the main constituent of the 2009;57(S1):161-177. extracts of abdominal tergite glands of virgin honeybee 2. Cowie I. New taxa and notes on Helicteres L. queens (Apis Mellifera L.), indicating its potential effect as (Malvaceae: ) from the Northern Territory, pheromone (Espelie et al., 1990; Trhlin and Rajchard, 2011; Australia. The Beagle, Records of the Museums and Art Villar et al., 2019) [34-36]. Galleries of the Northern Territory 2011;27:27-54. Other fatty acid derivatives were identified by GC-MS, such 3. Fernandes D, de Assis E, Souza M, Vanderlei P, as methyl and ethyl palmitate; phenolic compounds, such as Vanderlei M. Helicteres L. Species (Malvaceae Sensu phthalates and benzophenone derivatives; some steroidal and Lato) as source of new drugs: a review. Química Nova acyclic terpenoids, and other hydrocarbons. Phthalate 2020;43(6):787-803. DOI: https://doi.org/10.21577/0100- derivatives had been commonly pointed as contaminants in 4042.20170533 the process of purification in natural product studies (Bhakuni 4. Bravo O, Sánchez-González A, de Nova J, Pavón N. and Rawat, 2005; Venditti, 2018) [37, 38]. DBP, DEHP and Composición y estructura arbórea y arbustiva de la other phthalate derivatives are known as toxic components, vegetación de la zona costera de Bahía de Banderas, especially on reproductive system (Pan et al., 2006) [39]. These Nayarit, México [Composition and structure of trees and compounds also could be absorbed from water and soil into shrubs of the vegetation of the coastal area of Bahía de the plant root and be accumulated for them, thus identification Banderas, Nayarit, Mexico]. Botanical Sciences of phthalates in plant extracts or essential oils could be a 2016;94(3):603-623. signal of environmental pollution (Manayi et al., 2014) [40]. DOI: https://doi.org/10.17129/botsci.461 However, other studies suggest that phthalates derivatives 5. Notario S, Retana O, Vargas J. Mantenimiento de fauna could occur naturally, which are different of synthetic ones in silvestre durante la temporada de secas mediante parcelas terms of the abundance of 14C and its bond structure, leading de manejo [Sustaining wildlife during dry season through to its varied activities in the biological system (Zhang et al., management plots]. Tropical and Subtropical 2018; Narayan 2020) [41, 42]. The compound di-2-ethylhexyl Agroecosystems 2020;23(1):15(12). phthalate (DEHP), identified in this study, has been isolated 6. Fernandes A, Oliveira A. Flora das cangas da Serra dos from several organisms that appear to biosynthesize it Carajás, Pará, Brasil: Malvaceae [Flora of the canga of naturally, such as Calotropis gigantea (plant), Streptomyces the Serra dos Carajás, Pará, Brazil: Malvaceae]. sp. (bacteria) and Penicillium janthinellum (fungi), showing Rodriguésia 2018;69(3):1237-1254. interesting antibacterial and antitumor activities (Ortiz and DOI: http://dx.doi.org/10.1590/2175-7860201869325 Sansinenea, 2018) [43]. For that, it is difficult to confirm if 7. Stavis V, Ribeiro P, Gardenal B. Estrutura de fragmento phthalates identified here have a natural origin or are de floresta estacional em área de ecótono, Mato Grosso contaminants, but their presence might mean, in the first case, do Sul [Strucuture of an seasonal forest fragment at an a potential source of bioactive compounds, and, in the second, ecotono area, Mato Grosso do Sul, Brazil]. Brazilian a possible pollution in the area of recollection of sample or in Journal of Development 2020;6(8):59197-59213. the experimental processes. DOI:10.34117/bjdv6n8-366 Steroidal constituents (22E, 24R)-stigmasta-4,22-dien-3-one 8. Angarita H, Sanmartín S, Mercado G. Nuevos registros and δ-stigmast-4-en-3-one have also been identified together corológicos para Sucre (Sanguaré-Colombia) [New in bioactive extracts of Cenchrus setigerus (Singariya et al., chorological record to Sucre (Sanguaré-Colombia)]. 2014) [44] and Boophone haemanthoides (Ibrakaw et al., 2021) Revista Colombiana de Ciencia Animal-RECIA [45]. 2014;6(2):327-334. 9. Sanmartín-Sierra D, Angarita-Hernández D, Mercado- Conclusions Gómez J. Estructura y composición florística del bosque Results confirm that Helicteres guazumifolia is a source of seco tropical de Sanguaré-Sucre (Colombia) [Structure several phytochemical compounds, especially lipids (fatty and Floristic Composition of Tropical Dry Forest of acids, fatty alcohols, waxes, sterols and isoprenoids), which Sanguaré-Sucre (Colombia)]. Ciencia en Desarrollo are widely distributed in and could be used by 2016;7(2):43-56. themselves due to their ecological interactions. Furthermore,

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